Role of TRPM1 (Melastatin1) in the Biology of Human Melanocytes

TRPM1（Melastatin1）在人类黑素细胞生物学中的作用

基本信息

批准号：
7769851
负责人：
Vijayasaradhi Setaluri
金额：
$ 32.34万
依托单位：
UNIVERSITY OF WISCONSIN-MADISON
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-03-01 至 2013-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7769851
关键词：
Address Agonist Alternative Splicing Anabolism Antibodies Biochemical Biogenesis Biological Biological Assay Biology Calcium Carrier Proteins Cations Cell surface Cells Chemicals Chromatin Color Complex Cutaneous Melanoma Cytoplasmic Granules Data Differentiation and Growth Disease Divalent Cations Down-Regulation Drug or chemical Tissue Distribution Environmental Risk Factor Event Family Gene Expression Regulation Gene Silencing Genes Genetic Polymorphism Genetic Transcription Goals Growth Hair Hereditary Disease Homeostasis Human Human Biology Image In Situ In Vitro Invertebrates Ion Channel Protein Kidney Knowledge Length Malignant - descriptor Mammals Melanins Melanoma Cell Membrane Membrane Proteins Messenger RNA Metastatic Melanoma Methods Molecular Monophenol Monooxygenase Movement Mus Muscle Tonus Mutation Neoplasm Metastasis Nociception Organism Pain Perception Phenotype Physiological Physiological Processes Pigmentation physiologic function Pigments Plasmids Play Post-Translational Protein Processing Predisposition Process Proliferating Protein Family Protein p53 Proteins Publishing RNA Reagent Regulation Role Skin Skin Cancer Skin Pigmentation Smell Perception Smooth Muscle Solar Energy Source Stimulus Structure-Activity Relationship TP53 gene Taste Perception Testing Tetanus Helper Peptide Tissues Ultraviolet Rays Vision base blood pressure regulation human TRPM1 protein in vivo interest keratinocyte melanocortin receptor melanocyte melanoma member microphthalmia-associated transcription factor mutant neonatal human novel penis foreskin polypeptide promoter public health relevance receptor small hairpin RNA transcription factor tumor progression uptake

项目摘要

DESCRIPTION (provided by applicant): The goal of studies proposed here is to understand the functions and regulation of the melanocyte-specific transient receptor potential (TRP) ion channel protein, melastatin. Mammalian TRP channel protein family consists of nearly 30 members. TRPs are critical for functions of wide ranging physiological process, where cellular divalent cations, specifically Ca2+ and Mg2+ play important roles. These processes include photoperception, thermosensation, taste perception, mechanosensation, pain perception, Mg2+ homeostasis in kidney, smooth muscle tone and blood pressure regulation. Melastatin (TRPM1), the founding member of a novel family of TRP channel proteins known as TRPMs, was originally identified as suppressor of mouse melanoma metastasis. Independently, we identified TRPM1 as a gene induced in growth arrested human melanoma cells. Tissue distribution and promoter regulation by microphthalmia transcription factor (MITF) confirmed TRPM1 as a melanocyte-restricted TRP. Functional studies using a partial length human TRPM1 protein in heterologous HEK293 cells suggested that it is a constitutively active Ca2+ channel. Although loss of TRPM1 expression (as assessed by in situ RNA hybridization) is thought to correlate with aggressiveness of cutaneous melanoma, the exact function(s) of TRPM1 and how its expression is regulated in melanocytes are not known. To address these issues, we generated TRPM1 expression (constitutive and tet-inducible) shRNA plasmids and antibody reagents. Preliminary data showed that in primary neonatal human foreskin melanocytes a) knockdown of TRPM1 by shRNA inhibits accumulation of melanin pigment, b) TRPM1 expression is up-regulated by inducers of melanocyte differentiation and down regulated by p53 and c) malignant melanocytes do not tolerate expression of full-lengthTRPM1. These data suggested that TRPM1 is involved in regulation of growth and differentiated functions of epidermal melanocytes. In this proposal we will a) characterize biosynthesis, posttranslational modification, subcellular localization and membrane orientation and establish structure-function relationship of TRPM1, b) investigate the role of TRPM1 in growth of normal and malignant melanocytes and pigmentation using stable shRNA knockdown strategy, and c) investigate mechanism of regulation of TRPM1. These studies on understanding TRPM1 functions are significant in the context of growing interest in functions of TRP channels, association of mutations in TRP genes with genetic disorders and the potential of targeting TRPs for treatment of diseases arising from their malfunction. PUBLIC HEALTH RELEVANCE Melanocytes are the source of skin and hair pigmentation. Genetic defects and environmental factors that alter melanocyte function cause pigmentary disorders including skin cancer melanoma. This proposal investigates the role of a unique calcium transporting protein in maintaining proper functioning of human skin melanocytes. Knowledge gained from these studies could help develop new therapies for pigmentary disorders.

描述（由申请人提供）：此处提出的研究的目的是了解黑素细胞特异性瞬态受体电位（TRP）离子通道蛋白Melastatin的功能和调节。哺乳动物TRP通道蛋白家族由近30个成员组成。 TRP对于范围广泛的生理过程的功能至关重要，其中细胞二价阳离子（特别是Ca2+和Mg2+）起着重要作用。这些过程包括光感感受，热敏度，味觉感知，机械感应，疼痛感知，肾脏中的MG2+稳态，平滑肌张力和血压调节。 Melastatin（TRPM1）是一种新型TRP通道蛋白家族的创始成员，称为TRPMS，最初被鉴定为小鼠黑色素瘤转移的抑制剂。独立地，我们将TRPM1鉴定为在人类黑色素瘤细胞生长中诱导的基因。通过微观心脏转录因子（MITF）的组织分布和启动子调节证实TRPM1是黑色素细胞限制的TRP。在异源HEK293细胞中使用部分长度的人类TRPM1蛋白的功能研究表明，它是一种组成型活性Ca2+通道。尽管认为TRPM1表达的丧失（如原位RNA杂交评估）被认为与皮肤黑色素瘤的侵略性相关，TRPM1的确切功能以及其在黑色素细胞中如何调节其表达的功能尚不清楚。为了解决这些问题，我们生成了TRPM1表达（组成型和TET诱导型）shRNA质粒和抗体试剂。初步数据表明，在原发性新生儿外科体黑色素细胞中a）ShRNA敲低TRPM1的抑制黑色素色素的积累，b）TRPM1表达在黑色素细胞分化的诱导剂中被抑制了TRPM1的表达，而P53和C）由P53和C）降低了PRPM1和C）由P53和C）由PP53和C）耐受性黑素细胞的表达不耐受。这些数据表明，TRPM1参与了表皮黑素细胞的生长和分化功能的调节。在这项建议中，我们将a）表征生物合成，翻译后修饰，亚细胞定位和膜取向，并建立TRPM1的结构 - 功能关系，b）研究TRPM1在正常和恶性黑色素细胞和色素化的生长中的作用，并使用稳定的SHRNA敲低策略以及c negumencation Innegument of trpm1。在对TRP通道功能，TRP基因与遗传疾病的突变关联以及针对TRP的潜在治疗疾病引起的疾病引起的疾病的潜力的情况下，了解TRPM1功能的这些研究很重要。公共卫生相关性黑素细胞是皮肤和毛发沉着的来源。改变黑素细胞功能的遗传缺陷和环境因素会引起色素疾病，包括皮肤癌黑色素瘤。该提案研究了独特的钙转运蛋白在维持人皮肤素细胞正常功能中的作用。从这些研究中获得的知识可以帮助开发出新的色素疾病疗法。